Evaluation of women's aging influence on sperm passage inside the fallopian tube using 3D dynamic mechanical modeling.

The fallopian tubes play an important role in human fertility by facilitating the spermatozoa passage to the oocyte as well as later actively facilitating the fertilized oocyte transportation to the uterus cavity. The fallopian tubes undergo changes involving biological, physical, and morphological processes due to women aging, which may impair fertility. Here, we have modelled fallopian tubes of women at different ages and evaluated the chances of normal and pathological sperm cells reaching the fertilization site, the ampulla. By utilizing a unique combination of simulative tools, we implemented dynamic three-dimensional (3D) detailed geometrical models of many normal and pathological sperm cells swimming together in 3D geometrical models of three fallopian tubes associated with different women's age groups. By tracking the sperm cell swim, we found that for all age groups, the number of normal sperm cells in the ampulla is the largest, compared with the pathological sperm cells. On the other hand, the number of normal sperm cells in the fertilization site decreases due to the morphological and mechanical changes that occur in the fallopian tube with age. Moreover, in older ages, the normal sperm cells swim with lower velocities and for shorter distances inside the ampulla toward the ovary. Thus, the changes that the human fallopian tube undergoes due to women's aging have a significant influence on the human sperm cell motility. Our model of sperm cell motility through the fallopian tube in relation to the woman's age morphological changes provides a new scope for the investigation and treatment of diseases and infertility cases associated with aging, as well as a potential personalized medicine tool for evaluating the chances of a natural fertilization per specific features of a man's sperm and a woman's reproductive system.

Oocyte retrieval in social fertility preservation: challenging traditional beliefs on gonadotropin dosing and follicular size.

OBJECTIVES: To investigate treatment approaches for fertility preservation patients, with a focus on timing of oocyte retrieval, and to determine whether their characteristics differ from those of other IVF patients. Additionally, to evaluate the significance of follicle size on triggering day in the context of fertility preservation. METHODS: This retrospective cohort study was conducted in a tertiary, university-affiliated medical center. It compared 140 matched patients undergoing social fertility preservation to 140 patients undergoing IVF treatment due to male factor infertility. RESULTS: Patients undergoing fertility preservation received a higher initial gonadotropin dose and had more oocytes retrieved than the control group. Within the fertility preservation cohort, a negative correlation was observed between the rate of large follicles and the number of retrieved oocytes. While there was no significant association between rate of large follicles and oocyte maturation rate in the entire group, age-stratified analysis revealed a negative relationship. Analysis revealed that although traditional treatment determinants such as follicular size and gonadotropin dosing were considered, peak estradiol levels were consistently identified as significant predictors of treatment outcomes. CONCLUSIONS: Physicians may modify treatments for fertility preservation, emphasizing a higher gonadotropin dosage to maximize oocyte retrieval. Elevated estradiol levels can serve as a real-time predictive marker for the number of mature oocytes. While treatment strategies can influence outcomes, intrinsic patient factors, particularly baseline ovarian function, remain crucial. These results challenge beliefs regarding the importance of larger follicles and suggest the need for a tailored approach, considering patient age and specific fertility preservation objectives.

Obstetric and maternal outcomes of IVF and oocyte donation pregnancies among women ages 40-45-a large cohort study.

PURPOSE: To analyze the perinatal and maternal outcomes of women ranging in age from 40 to 45 years who gave birth after in vitro fertilization or oocyte donation, compared to spontaneous conception. METHODS: This retrospective cohort study used electronic data from a national healthcare service from 2000 through 2019. Three groups were compared: spontaneous pregnancy (SC), in vitro fertilization (IVF) utilizing autologous oocytes, and pregnancies resulting from oocyte donation (OD). The primary study outcomes were preterm labor (PTL) before 37 weeks of gestation, and infants classified as small for gestational age (SGA). RESULTS: The cohort included 26,379 SC, 2237 IVF pregnancies, and 300 OD pregnancies for women ages 40-45 years at delivery. Women with OD or IVF had a higher incidence of PTL < 37 weeks compared to women with SC (19.7% vs. 18% vs. 6.9%, p = 0.001), PTL < 34 (7% vs. 4.5% vs. 1.4%, p = 0.001), PTL < 32 (3.7 vs. 2.1 vs. 0.6, p = 0.001). A multivariable logistic regression for PTL < 37 weeks demonstrated that age (OR = 1.18) and hypertensive diseases (OR = 3.4) were statistically significant factors. The OD group had a lower rate of SGA compared to SC (1% vs. 4.3%, p = 0.001), while the IVF group had a higher rate of SGA compared to SC (9.1% vs. 4.3%, p = 0.001). Hypertensive diseases in pregnancy were significantly higher among the OD group and the IVF group compared to SP pregnancies (3.3% vs. 1%, p = 0.002; 2.3% vs. 1%, p = 0.001, respectively). CONCLUSIONS: Women ages 40-45 undergoing IVF or OD have a greater risk of PTL, possibly due to higher rates of hypertensive disorders of pregnancy.

Stain-Free Sperm Analysis and Selection for Intracytoplasmic Sperm Injection Complying with WHO Strict Normal Criteria.

This multi-center study evaluated a novel microscope system capable of quantitative phase microscopy (QPM) for label-free sperm-cell selection for intracytoplasmic sperm injection (ICSI). Seventy-three patients were enrolled in four in vitro fertilization (IVF) units, where senior embryologists were asked to select 11 apparently normal and 11 overtly abnormal sperm cells, in accordance with current clinical practice, using a micromanipulator and 60× bright field microscopy. Following sperm selection and imaging via QPM, the individual sperm cell was chemically stained per World Health Organization (WHO) 2021 protocols and imaged via bright field microscopy for subsequent manual measurements by embryologists who were blinded to the QPM measurements. A comparison of the two modalities resulted in mean differences of 0.18 µm (CI -0.442-0.808 µm, 95%, STD-0.32 µm) for head length, -0.26 µm (CI -0.86-0.33 µm, 95%, STD-0.29 µm) for head width, 0.17 (CI -0.12-0.478, 95%, STD-0.15) for length-width ratio and 5.7 for acrosome-head area ratio (CI -12.81-24.33, 95%, STD-9.6). The repeatability of the measurements was significantly higher in the QPM modality. Surprisingly, only 19% of the subjectively pre-selected normal cells were found to be normal according to the WHO2021 criteria. The measurements of cells imaged stain-free through QPM were found to be in good agreement with the measurements performed on the reference method of stained cells imaged through bright field microscopy. QPM is non-toxic and non-invasive and can improve the clinical effectiveness of ICSI by choosing sperm cells that meet the strict criteria of the WHO2021.

Dynamic 3D Modeling for Human Sperm Motility through the Female Cervical Canal and Uterine Cavity to Predict Sperm Chance of Reaching the Oocyte.

Sperm motility in the female genital tract is a key factor in the natural selection of competent cells that will produce a healthy offspring. We created a dynamic three-dimensional (3D) mechanical model of human sperm cells swimming inside cervical canal and uterine cavity dynamic 3D models, all generated based on experimental studies. Using these simulations, we described the sperm cells' behaviors during swimming inside the 3D tract model as a function of 3D displacement and time. We evaluated normal- and abnormal-morphology sperm cells according to their chances of reaching the oocyte site. As expected, we verified that the number of normal sperm cells that succeeded in reaching the fallopian tube sites is greater than the number of abnormal sperm cells. However, interestingly, after inspecting various abnormal sperm cells, we found out that their scores changed compared to swimming in an infinite medium, as is the case with in vitro fertilization. Thus, the interactions of abnormal sperm cells and the complicated geometry and dynamics of the uterus are significant factors in the filtering of abnormal sperm cells until they reach the oocyte site. Our study provides an advanced tool for sperm analysis and selection criteria for fertility treatments.

Sperm-cell DNA fragmentation prediction using label-free quantitative phase imaging and deep learning.

In intracytoplasmic sperm injection (ICSI), a single sperm cell is selected and injected into an egg. The quality of the chosen sperm and specifically its DNA fragmentation have a significant effect on the fertilization success rate. However, there is no method today to measure the DNA fragmentation of live and unstained cells during ICSI. We present a new method to predict the DNA fragmentation of sperm cells using multi-layer stain-free imaging data, including quantitative phase imaging, and lightweight deep learning architectures. The DNA fragmentation ground truth is achieved by staining the cells with acridine orange and imaging them via fluorescence microscopy. Our prediction model is based on the MobileNet convolutional neural network architecture combined with confidence measurement determined by distances between vectors in the latent space. Our results show that the mean absolute error for cells with high prediction confidence is 0.05 and the 90th percentile mean absolute error is 0.1, where the range of DNA fragmentation score is [0,1]. In the future, this model may be applied to improve cell selection by embryologists during ICSI.

Pretreatment with gonadotropin-releasing hormone antagonist protects against chemotherapy-induced testicular damage 'in mice.

Background: Testicular toxicity following chemotherapy is of increasing importance with the continuous improvement of survival rates. Gonadotropin-releasing hormone (GnRH) was suggested to protect testis against such toxicity; however, its suppressive quality and mechanism of action are still unclear. We examined whether and how pretreatment with GnRH antagonist protects against the testicular damage caused by chemotherapy. Methods: Mature male mice were injected subcutaneously eight times in 2-day intervals with either saline or GnRH antagonist (Cetrotide; 1 g/mg), followed by an intraperitoneal injection with either saline or cyclophosphamide (CTX;100 mg/kg BW) and sacrificed 2 weeks or 3 months later. Testicular weight, epididymis weight, epididymal sperm count and sperm motility were measured. Serum anti-Müllerian hormone (AMH) was measured by enzyme-linked immunosorbent assay. Immunohistochemistry (Ki-67), immunofluorescence (PCNA, CD34), terminal transferase-mediated deoxyuridine 5-triphosphate nick-end labeling (TUNEL) and computerized analysis were performed to examine testicular proliferation, apoptosis and vascularization. Quantitative real-time PCR was used to assess the amount of spermatogonial reserve (Id4 and Gfra1 mRNAs). Results: Pretreatment with GnRH antagonist transiently reduced testicular weight, epididymal weight, germinal proliferation and sperm count; it also abolished the permanent long-term effect of CTX on these parameters and prevented cyclophosphamide-induced testicular toxicity characterized by apoptosis and serum AMH increase and irreversible loss of spermatogonial reserve. Conclusions: Our findings imply that pretreatment with GnRH antagonist temporarily reduces spermatogenesis and may be used as pretreatment for reducing chemotherapeutic testicular toxicity.

Irinotecan (CPT-11) Treatment Induces Mild Gonadotoxicity.

Background: Gonadal toxicity following chemotherapy is an important issue among the population of young cancer survivors. The inhibitor of DNA topoisomerase I, irinotecan (CPT-11), is widely used for several cancer types. However, little is known about the effect of irinotecan on the fertility of both genders. Thus, the aim of the present study was to evaluate irinotecan gonadotoxicity, using a mouse model. Methods: Mature male and female mice were injected intraperitoneally with either saline (), irinotecan (100 mg/kg) or cyclophosphamide (100 mg/kg); and sacrificed one week or three months later for an acute or long-term toxicity assessment, respectively. We used thorough and advanced fertility assessment by already established methods: Gonadal and epididymal weights, as well as sperm count and sperm motility were determined; serum anti-Müllerian hormone (AMH) was measured by ELISA. Immunohistochemistry (Ki-67), immunofluorescence (PCNA, CD34), terminal transferase-mediated deoxyuridine 5-triphosphate nick-end labeling (TUNEL) and computerized analysis were performed to examine gonadal proliferation, apoptosis and vascularization. qPCR was used to assess the amount of testicular spermatogonia (Id4 and Gafra1 mRNA) and ovarian primordial oocytes reserves (Sohlh2, Nobox and Figla mRNA). Results: Females: Irinotecan administration induced acute ovarian apoptosis and decreased vascularity, as well as a mild, statistically significant, long-term decrease in the number of growing follicles, ovarian weight, and ovarian reserve. Males: Irinotecan administration caused an acute testicular apoptosis and reduced testicular spermatogenesis, but had no effect on vascularity. Irinotecan induced long-term decrease of testicular weight, sperm count and testicular spermatogonia and caused elevated serum AMH. Conclusion: Our findings imply a mild, though irreversible effect of irinotecan on mice gonads.

Tumor Lymphocyte Infiltration Is Correlated with a Favorable Tumor Regression Grade after Neoadjuvant Treatment for Esophageal Adenocarcinoma.

(1) Background: We aimed to explore the association between neoadjuvant treatment, tumor-infiltrating immune lymphocyte (TIL), and tumor-associated macrophage (TAM) and survival in patients with esophageal adenocarcinoma. (2) Methods: Patients who underwent esophagectomy were divided into three groups according to their treatment modality and tumor regression grade (TRG): (i) surgery-only group (SG), (ii) good responders (GR) group (TRG 0−1), and (iii) bad responders (BR) group (TRG 2−3). We then carried out statistical correlations of the immunofluorescence analysis of the immune infiltrate in the esophageal surgical specimens with several clinical and pathological parameters. In addition, we analyzed The Cancer Genomic Atlas (TCGA) dataset for differences in TILs, TAMs, and protein expression in immune pathways. (3) Results: Forty-three patients (SG­15, GR­13, and BR­13) were evaluated. The highest enrichment of CD3+ (p < 0.001), CD8+ (p = 0.001) and CD4+ (p = 0.009) was observed in the stroma of GR patients. On multivariate analysis, only CD8+ T cell and signet-ring features were independent prognostic factors for overall survival. In TCGA analysis, we identified overexpression of TAM and colony-stimulating factor 1 receptor (CSF-1R). (4) Conclusions: High enrichment of lymphocyte subpopulations in the microenvironment of esophageal adenocarcinoma is associated with a favorable response to neoadjuvant treatment and an improved patient outcome.

Prediction of Sperm Progression in Three Dimensions Using Rapid Optical Imaging and Dynamic Mechanical Modeling.

We present a multidisciplinary approach for predicting how sperm cells with various morphologies swim in three-dimensions (3D), from milliseconds to much longer time scales at spatial resolutions of less than half a micron. We created the sperm 3D geometry and built a numerical mechanical model using the experimentally acquired dynamic 3D refractive-index profiles of sperm cells swimming in vitro as imaged by high-resolution optical diffraction tomography. By controlling parameters in the model, such as the size and shape of the sperm head and tail, we can then predict how different sperm cells, normal or abnormal, would swim in 3D, in the short or long term. We quantified various 3D structural factor effects on the sperm long-term motility. We found that some abnormal sperm cells swim faster than normal sperm cells, in contrast to the commonly used sperm selection assumption during in vitro fertilization (IVF), according to which sperm cells should mainly be chosen based on their progressive motion. We thus establish a new tool for sperm analysis and male-infertility diagnosis, as well as sperm selection criteria for fertility treatments.

Anti-HER2/neu Antibody Reduces Chemotherapy-Induced Ovarian Toxicity-From Bench to Bedside.

BACKGROUND: Trastuzumab, a humanized anti-human epidermal growth factor receptor 2 (HER2/neu) antibody, is considered a standard treatment in addition to chemotherapy in the adjuvant setting for HER2/neu-positive breast cancer, yet its impact on fertility and ovarian reserve remains obscure. We aimed to study the effect of anti-HER2/neu on chemotherapy-induced ovarian toxicity in both clinical and preclinical settings. METHODS: We prospectively enrolled breast cancer patients below the age of 42 years who were treated with chemotherapy with or without trastuzumab into the study. Anti-Müllerian hormone (AMH) was measured 6 and 12 months post-chemotherapy as an ovarian reserve indicator. In the animal model, pubertal mice were injected with cyclophosphamide or paclitaxel with or without anti-HER2/neu, or saline, and sacrificed 1 week or 3 months later. Ovarian apoptosis, proliferation and vascularity were measured by immunohistochemistry and ovarian reserve was measured by morphometric analysis and serum-AMH. RESULTS: Thirty-three patients with early breast cancer were enrolled into the study. Nineteen patients had HER2/neu negative cancer and were treated with chemotherapy and 14 had HER2/neu positive cancer and were treated with chemotherapy and trastuzumab. In all patients, AMH levels declined to undetectable values immediately post-treatment, but regained for 57.1% of the HER2/neu positive cohort and 36.8% of the negative cohort (p < 0.05). In the preclinical setting, anti-HER2/neu antibody, in combination with chemotherapy, displayed lessened ovarian and vascular damage. CONCLUSIONS: Our results indicate that trastuzumab may alleviate chemotherapy-induced ovarian toxicity that may be mediated via its effect on ovarian vasculature.

Cancer During Pregnancy: The Role of Vascular Toxicity in Chemotherapy-Induced Placental Toxicity.

Breast cancer is diagnosed in ~0.3% of pregnant women. Studies that have addressed gestational and neonatal outcomes of chemotherapy during pregnancy have demonstrated increased gestational complications including preeclampsia and intrauterine growth retardation. We hypothesized that anthracycline-induced gestational complications could be derived from direct toxicity on the placenta vasculature. Pregnant ICR mice (day E12.5) were treated with doxorubicin (DXR; 8 mg/kg) or saline, while their umbilical cord blood flow was imaged by pulse-wave (PW) Doppler. Mice were euthanized on day E18.5, and their embryos and placentae were collected for further analysis. Unlike control mice, the DXR-treated mice presented an acute change in the umbilical cord's blood flow parameters (velocity time integral and heart rate interval), reduced embryos' weight, reduced placenta efficiency, and modulation in vascular-related pathways of treated placenta proteomics. Apoptosis and proliferation were also enhanced, as demonstrated by TUNEL and proliferating cell nuclear antigen (PCNA) analysis. We further examined the placentae of patients treated with epirubicin (EPI), who had been diagnosed with breast cancer during pregnancy (weeks 27-35). The immunohistochemistry of the EPI-treated human placentae showed enhanced proliferation and apoptosis as compared with matched chemo-naïve placentae, as well as reduced neovascularization (CD34). Our findings suggest that anthracycline-induced vascular insult promotes placental toxicity, and could point to potential agents designated to offset the damage and to reduce gestational complications in pregnant cancer patients.

Holographic virtual staining of individual biological cells.

Many medical and biological protocols for analyzing individual biological cells involve morphological evaluation based on cell staining, designed to enhance imaging contrast and enable clinicians and biologists to differentiate between various cell organelles. However, cell staining is not always allowed in certain medical procedures. In other cases, staining may be time-consuming or expensive to implement. Staining protocols may be operator-sensitive, and hence may lead to varying analytical results, as well as cause artificial imaging artifacts or false heterogeneity. We present a deep-learning approach, called HoloStain, which converts images of isolated biological cells acquired without staining by holographic microscopy to their virtually stained images. We demonstrate this approach for human sperm cells, as there is a well-established protocol and global standardization for characterizing the morphology of stained human sperm cells for fertility evaluation, but, on the other hand, staining might be cytotoxic and thus is not allowed during human in vitro fertilization (IVF). After a training process, the deep neural network can take images of unseen sperm cells retrieved from holograms acquired without staining and convert them to their stainlike images. We obtained a fivefold recall improvement in the analysis results, demonstrating the advantage of using virtual staining for sperm cell analysis. With the introduction of simple holographic imaging methods in clinical settings, the proposed method has a great potential to become a common practice in human IVF procedures, as well as to significantly simplify and radically change other cell analyses and techniques such as imaging flow cytometry.

Stain-free interferometric phase microscopy correlation with DNA fragmentation stain in human spermatozoa.

Acridine orange (AO) staining is used to diagnose DNA fragmentation status in sperm cells. Interferometric phase microscopy (IPM) is an optical imaging method based on digital holographic microscopy that provides quantitative morphological and refractive index imaging of cells in vitro without the need for staining. We have imaged sperm cells using stain-free IPM in order to estimate different cellular parameters, such as acrosome dry mass and size, in addition to an embryologist evaluation according to the World Health Organization (WHO)-2010 criteria. Following this, the same sperm cells were stained by AO, imaged using a fluorescence confocal microscope and assessed by the AO-emitted color, forming five DNA fragmentation groups. These DNA fragmentation groups were correlated with the embryologist-based classification and the IPM-based morphological parameters. Our results indicate on significant differences in the IPM-based parameters between groups with different fragmentation levels. Based on the validation with AO, we conclude that stain-free IPM images analyzed digitally may assist in selecting sperm cells with intact DNA prior to intracytoplasmic sperm injection. This information may potentially increase percentage of successful pregnancies.

Treosulfan induces distinctive gonadal toxicity compared with busulfan.

Treosulfan (L-treitol-1,4-bis-methanesulfonate) has been increasingly incorporated as a main conditioning protocol for hematopoietic stem cell transplantation in pediatric malignant and non-malignant diseases. Treosulfan presents lower toxicity profile than other conventional alkylating agents containing myeloablative and immunosuppressive traits such as busulfan. Yet, whereas busulfan is considered highly gonadotoxic, the gonadal toxicity profile of treosulfan remains to be elucidated. To study the gonadotoxicity of treosulfan, pubertal and prepubertal male and female mice were injected with treosulfan or busulfan and sacrificed one week, one month or six months later. Testicular function was assessed by measurements of sperm properties, testes and epididymides weights as well as markers for testicular reserve, proliferation and apoptosis. Ovarian function was assessed by measurements of ovary weight and markers for ovarian reserve, proliferation and apoptosis. Treosulfan testicular toxicity was milder than that of busulfan toxicity; possibly by sparing the stem spermatogonia in the testicular sanctuary. By contrast, ovarian toxicity of both treosulfan and busulfan was severe and permanent and displayed irreversible reduction of reserve primordial follicles in the ovaries. Our data indicate that treosulfan exerts a different gonadal toxicity profile from busulfan, manifested by mild testicular toxicity and severe ovarian toxicity.

Premature ovarian aging in BRCA carriers: a prototype of systemic precocious aging?

PURPOSE: Though former evidence implies a correlation of breast cancer susceptibility gene (BRCA) mutation with reduced ovarian reserve, the data is yet inconsistent. Our aim was to investigate biomarkers of ovarian aging in a cohort of young healthy carriers of the BRCA mutation. We hypothesized that the role played by BRCA genes in aging pathways is not exclusive to the ovary. EXPERIMENTAL DESIGN: Healthy female BRCA carriers, 40 years or younger and healthy male BRCA carriers, 50 years or younger, were enrolled in the study. Serum anti-mullerian Hormone (AMH), fibroblast growth factor-23 (FGF-23), Klotho and IL-1 were measured by enzyme-linked immunosorbent assay (ELISA). Ovarian AMH and protein kinase B (AKT) mRNA from BRCA carriers who underwent prophylactic oophorectomy and from age-matched, healthy, non-carriers who underwent partial oophorectomy due to benign conditions were analyzed by qPCR. RESULTS: Thirty-three female (median age 35y) and 20 male (44y) BRCA carriers were enrolled into the study and matched to control non-carriers (34y and 43y, respectively). Serum AMH level was significantly lower in BRCA female carriers than in both non-carrier controls and age-matched nomograms. The levels of ovarian AMH and AKT mRNA were significantly lower in carriers than in controls. The systemic aging cytokines FGF-23, klotho and IL-1 displayed a differential expression in carriers of both genders. FGF-23 level was higher in carriers (P=0.06). CONCLUSIONS: Our results suggest a link between BRCA mutation, accelerated ovarian aging and systemic aging-related pathophysiology.

Individual sperm selection by microfluidics integrated with interferometric phase microscopy.

The selection of sperm cells possessing normal morphology and motility is crucial for many assisted reproductive technologies (ART), especially for intracytoplasmic sperm injection (ICSI), as sperm quality directly affects the probability of inducing healthy pregnancy. We present a novel platform for real-time quantitative analysis and selection of individual sperm cells without staining. Towards this end, we developed an integrated approach, combining interferometric phase microscopy (IPM), for stain-free sperm imaging and real-time automatic analysis based on the sperm cell 3D morphology and contents, with a disposable microfluidic device, for sperm selection and enrichment. On testing the capabilities of the microfluidic device, we obtained successful selection of sperm cells with a selectivity of 89.5±3.5%, with no negative-decision sperm cells being inadvertently selected. In addition, we demonstrate the accuracy of sperm cell analysis using IPM by comparing the quantitative analysis produced by our IPM-based algorithm to the qualitative visual analysis performed independently by an experienced embryologist, which resulted in precision and specificity of 100%. We believe that the presented integrated approach has the potential to dramatically change the way sperm cells are selected for ICSI and other ART procedures, making the selection process more objective, quantitative and automatic, and thereby increasing success rates.

Can molecular profiling enhance radiotherapy? Impact of personalized targeted gold nanoparticles on radiosensitivity and imaging of adenoid cystic carcinoma.

Personalized molecular profiling has an established role in selection of treatment for metastatic disease; however, its role in improving radiosensitivity and functional imaging has not been evaluated. In the current study, we examined molecular profiling as a tool for designing personalized targeted gold nanoparticles (GNP) to serve as dual-modal tumor radiosensitizers and functional imaging enhancers. To this end, molecular profiling of a patient's salivary gland adenoid cystic carcinoma (ACC) was performed, and anaplastic lymphoma kinase (ALK) mutation was detected. The extracted tumor was subcutaneously injected into mice, which were then treated either with radiation, the specific ALK inhibitor crizotinib, or a combination of therapies. One of these combinations, namely, ALK-targeted GNP (via crizotinib coating), was found to enhance radiation treatment, as demonstrated by a significant decrease in tumor volume over 24 days. In parallel, ALK-targeted GNP substantially augmented tumor visualization via computed tomography. The mechanism of radiosensitivity enhancement was mostly related to a diminished cell repair mechanism in tumors, as demonstrated by proliferating cell nuclear antigen staining. These findings indicate that personalized molecular profiling is an effective technique for enhancing cancer theranostics.

Automated analysis of individual sperm cells using stain-free interferometric phase microscopy and machine learning.

Currently, the delicate process of selecting sperm cells to be used for in vitro fertilization (IVF) is still based on the subjective, qualitative analysis of experienced clinicians using non-quantitative optical microscopy techniques. In this work, a method was developed for the automated analysis of sperm cells based on the quantitative phase maps acquired through use of interferometric phase microscopy (IPM). Over 1,400 human sperm cells from 8 donors were imaged using IPM, and an algorithm was designed to digitally isolate sperm cell heads from the quantitative phase maps while taking into consideration both the cell 3D morphology and contents, as well as acquire features describing sperm head morphology. A subset of these features was used to train a support vector machine (SVM) classifier to automatically classify sperm of good and bad morphology. The SVM achieves an area under the receiver operating characteristic curve of 88.59% and an area under the precision-recall curve of 88.67%, as well as precisions of 90% or higher. We believe that our automatic analysis can become the basis for objective and automatic sperm cell selection in IVF. © 2017 International Society for Advancement of Cytometry.

Regulation of GVBD in mouse oocytes by miR-125a-3p and Fyn kinase through modulation of actin filaments.

Meiotically arrested oocytes are characterized by the presence of the nuclear structure known as germinal-vesicle (GV), the breakdown of which (GVBD) is associated with resumption of meiosis. Fyn is a pivotal factor in resumption of the first meiotic division; its inhibition markedly decreases the fraction of oocytes undergoing GVBD. Here, we reveal that in mouse oocytes Fyn is post-transcriptionally regulated by miR-125a-3p. We demonstrate that in oocytes resuming meiosis miR-125a-3p and Fyn exhibit a reciprocal expression pattern; miR-125a-3p decreases alongside with an increase in Fyn expression. Microinjection of miR-125a-3p inhibits GVBD, an effect that is markedly reduced by Fyn over-expression, and impairs the organization of the actin rim surrounding the nucleus. Lower rate of GVBD is also observed in oocytes exposed to cytochalasin-D or blebbistatin, which interfere with actin polymerization and contractility of actin bundles, respectively. By down-regulating Fyn in HEK-293T cells, miR-125a-3p reduces the interaction between actin and A-type lamins, which constitute the nuclear-lamina. Our findings suggest a mechanism, by which a decrease in miR-125a-3p during oocyte maturation facilitates GVBD by allowing Fyn up-regulation and the resulting stabilization of the interaction between actin and A-type lamins.